Nitric oxide synthase (NOS) catalyzes the oxidation of L-arginine to nitric oxide (NO) and citrulline; NADPH and O.sub.2 are cosubstrates. Three major isoforms of NOS have been identified to date. The neuronal (nNOS) and endothelial (eNOS) isoforms are constitutively expressed and are regulated by Ca.sup.2+ /calmodulin, whereas the activity of the inducible isoform (iNOS) is controlled transcriptionally and is not affected by changes in intracellular Ca.sup.2+. The three isoforms are all comprised of a C-terminal reductase domain that binds NADPH and the cofactors FAD and FMN and an N-terminal oxygenase domain that binds L-arginine and heme and tetrahydrobiopterin cofactors.
Nitric oxide synthase-derived NO is important in many physiological processes including blood pressure homeostasis, neurotransmission and immune function, but overproduction of NO can have pathological consequences. For example, excess NO resulting from overexpression of iNOS in response to endotoxin or inflammatory cytokines is a major contributor to the vascular disregulation seen in septic shock and in patients receiving interleukin-2-based immunotherapy. Inappropriate activation of nNOS is implicated in chronic visceral pain, in migraine headache, and in several neurodegenerative diseases (e.g., Parkinson's disease) and contributes to post-ischemic reperfusion injury in stroke.
Because NO has beneficial effects, there is great interest in NOS inhibitors that preferentially inhibit only the NOS isoform causing the particular pathology being treated. For example, iNOS specific inhibitors are of interest as drugs for the treatment of a variety of inflammatory and autoimmune conditions. Inhibitors that preferentially inhibit nNOS are of interest for the treatment of migraine headaches, mild to moderate pain due to the stimulation of nNOS-containing neurons including chronic visceral pain, stroke, Parkinson's disease, schizophrenia, drug abuse and epilepsy.
It has been shown that the S-alkyl-L-thiocitrullines show modest selectivity (up to 50-fold) for nNOS over eNOS and iNOS, and it has been proposed that these compounds may be of use in treating disorders involving overstimulation of nNOS (e.g., stroke). Improved potency and isoform selectivity would, however, be highly desirable. Moreover, the S-alkyl-L-thiocitrullines are reversible inhibitors of nNOS, so they would have to be administered repetitively in successive doses. An irreversible inhibitor of nNOS would have the advantage of requiring only a few or a single administration(s).